To date the turnover of lymphocyte populations in humans has not been systematically examined. Studies of the CD4+ T-cell rebound following effective antiretroviral therapy have suggested a rapid lymphocyte turnover. However, that conclusion was based on the assumption that the cell increase was due to regeneration rather than redistribution. Moreover, a study on telemore lengths of CD4+ T-cells in HIV-1-infected persons had suggested that there was no increased turnover of the CD4+ lymphocyte population, contradicting a large volume of literature describing increased evidence of lymphocyte activation and apoptosis associated with HIV-1 infection. To address this controversy, we recently completed a direct labeling study using bromodeoxyuridine to mark proliferating cells in normal and SIV-infected macaques. Indeed heightened turnover was observed in SIV infection for CD4+ T-cells as well as for other cell populations. We now plan to extend this type of labeling study to humans using a newly developed technique that is clinically safe. We will use glucose made with the stable isotope deuterium at the 6,6-position ("deuterated glucose") to label newly synthesized DNA within proliferating cells. The extent of the cellular proliferation could be quantified by isolating genomic DNA from the cell population of interest (CD4+ and CD8+ memory and nave cells; NK and B lymphocytes) to measure the degree of isotopic cont... enrichment in purine deoxyribonucleosides (dA or dG) with the use of mass spectrometry. In this regard, Julio Padovan will help us perform mass spectrometry to measure isotope incorporation in a cross-sectional study to directly define lymphocyte kinetics in normal individuals, as well as in HIV-1-infected persons who are clinically stable and untreated. In addition, we shall repeat the labeling studies in the infected cases after HIV-1 replication has been potently suppressed by combination antiretroviral therapy. These studies should provide valuable insights into human lymphocyte homeostasis in general, as well as into the pathogenic processes involved in lymphocyte destruction in HIV-1 infection in particular.